It is known that latent and chronically persistent viral infections can be activated or reactivated by immunosuppression, or conversely that the immune system suppresses acute diseases which may be caused by a latent virus (for example a latent herpes virus infection recurs as a result of immunosuppression in the form of lip vesicles in cases of stress or the administration of cortisone). It is also known that chronically persistent latent viral infections can only be treated with difficulty or not at all using conventional low-molecular-weight antiviral substances.
It was demonstrated that class I restricted cytotoxic T cells were capable of inhibiting hepatocellular HBV gene expression in HBV-transgenic mice, and that this process was caused by TNF-α and IFN-γ.
It is also known that in the case of chronically persistent viral infections a superinfection with another virus can produce antiviral effects against the chronically persistent virus. The dependence of this effect on interferons such as IFN-γ, as well as other cytokines and chemokines, such as TNF-α, which are secreted by T cells, natural killer cells and macrophages, has been demonstrated.
BAYPAMUN®, a pharmaceutical product for inducing “paraspecific immunity”, i.e., a pharmaceutical product for inducing the unspecific immune system, is used therapeutically, metaphylactically and prophylactically for the treatment of animals in need. BAYPAMUN® is manufactured from chemically inactivated PPVO strain D1701 (see German Patent DE3504940). The inactivated PPVO induces in animals non-specific protection against infections with the most diverse types of pathogens. It is assumed that this protection is mediated via various mechanisms in the body's own defense system. These mechanisms include the induction of interferons, the activation of natural killer cells, the induction of “colony-stimulating activity” (CSA) and the stimulation of lymphocyte proliferation. Earlier investigations of the mechanism of action demonstrated the stimulation of interleukin-2 and interferon-α.
The processes for the production of the above-mentioned pharmaceutical compositions are based on the replication of the virus in cultures of suitable host cells.
One aspect of the invention relates to the use of particle-like structures comprising recombinant proteins of the invention. These particle-like structures can be, e.g., fusion proteins, protein-coated particles or virus-like particles.
Methods to produce fusion proteins, protein-coated particles or virus-like particles comprising recombinant proteins of the invention are well known to persons skilled in the art: Casal (Biotechnol. Genet. Eng. Rev. (2001) 18:73-87) describes the use of baculovirus expression systems for the generation of virus-like particles. Ellis (Curr. Opin. Biotechnol. (1996) 7(6):646-652) presents methods to produce virus-like particles and the application of suitable adjuvants. Roy (Intervirology (1996) 39(1-2):62-71) presents genetically engineered particulate virus-like structures and their use as vaccine delivery systems. Methods to produce fusion proteins are also well known to the person skilled in the art (Gaudin, et al., Gen. Virol. (1995) 76:1541-1556; Hughson, Curr. Biol. (1995) 5(3):365-374; Uhlen, et al., Curr. Opin. Biotechnol. (1992) 3(4):363-369). Known to the person skilled in the art is also the preparation of protein-coated micro- and nanospheres (Arshady, Biomaterials (1993) 14(1):5-15). Proteins can be attached to biodegradable microspheres (Cleland, Pharm Biotechnol. (1997) 10:1-43) or attached to other polymer microspheres (Hanes, et al., Pharm. Biotechnol. (1995) 6:389-412) such as, e.g., polysaccharides (Janes, et al., Adv. Drug Deliv. Rev. (2001) 47(1):83-97).
PPVO NZ2 is another Parapoxvirus strain that exhibits immunostimulatory effects when administered in inactivated form to mammals.
The closest prior art describes the construction of an expression library representing about 95% of the PPVO NZ2 genome using the Vaccina lister virus to create recombinant viruses comprising the complete Vaccina lister genome and various fragments of the PPVO genome (Mercer, et al., Virology, (1997) 229:193-200). For the construction of the library, 16 PPVO DNA fragments with an average size of 11.4 kb were inserted into the Vaccinia lister genome. Each fragment was mapped relative to the PPVO restriction endonuclease maps but was otherwise uncharacterized (FIG. 1). It was found that a major portion of the PPVO genes were expressed in cells infected by the recombinant virus. The authors also showed that the entirety of all PPVO proteins expressed by some of the recombinant viruses of the expression library was able to provide protection against challenge with virulent PPVO. Expression of PPVO genes of the individual recombinant viruses has been demonstrated by immunofluorescence and immune precipitation (Mercer, et al., Virology (1997) 229:193-200).
To identify components of PPVO responsible for the vaccinating activity of PPVO, the Vaccinia lister/PPVO NZ2 expression library was applied.
Based on the above background it was desirable to develop PPVO based pharmaceutical compositions with antiviral and anti-tumor efficacy as well as with efficacy in paraimmunization and other desirable therapeutic effects. It was also desirable to obtain a pharmaceutical composition that exerts its full therapeutic effect while showing fewer side effects. It was furthermore desirable to find methods to produce PPVO based pharmaceutical compositions in large quantities and in economically advantageous manners.
These desirable effects have been achieved by the systematic use of selected recombinant proteins of PPVO alone or in combination with other recombinant proteins from PPVO for the preparation of pharmaceutical compositions for the treatment of objects in need.